Down syndrome is caused by trisomy 21 (TS21) (an extra copy of chromosome 21). TS21 leads to a broad spectrum of phenotypes including mental retardation and various heart defects. TS21 is the primary model for understanding aneuploidies or aberrant chromosomal numbers and the numerous genetic disorders they cause. The most widely accepted hypothesis concerning the pathology of Down syndrome is known as the gene dosage hypothesis. Chromosome 21 RNA transcripts are increased in individuals with Down syndrome as compared to the RNA transcripts in normal individuals. Many transcripts are expressed at much higher or lower levels than the theoretical 1.5 fold increase, which indicates that factors other than the three copies of chromosome 21 are involved in regulating the RNA transcript levels. Epigenetic modifications are a major source of transcription control. The broad, long-term objective of the proposed research is to understand any epigenetic changes occurring in TS21 as a model for other disorders caused by increases in chromosomal copy number. Epigenetic changes will be examined by determining the methylation status of CpG islands using bisulfite sequencing. Advances in bisulfite sequencing technology will allow large amounts of data to be generated, providing the opportunity for comprehensive data on methylation changes. Data analysis and visualization tools will be developed to provide an accurate and complete understanding of the data. Abnormal amounts of genetic material are a primary cause of genetic disorders. In Down syndrome, an extra copy of chromosome 21 leads to an increase in the level of RNA generated from chromosome 21 and ultimately the features of the disorder. Understanding modification of the DNA (which plays a significant role in how much RNA is generated) is an important step in understanding these types of genetic disorders. The proposed research will examine DNA modifications and their relationship to RNA levels in Down syndrome. The results of this research will act as a model for other disorders caused by abnormal amounts of genetic material.